FIGURE 4.

Role of PAS2 in TodS/TodT signal transduction. A, SEC analysis of purified PAS2(611–729). Eluted PAS2 protein was compared with the molecular weight standard marker carbonic anhydrase (29 kDa) and analyzed as a dimer in solution. B, the modeled PAS2 structure (magenta) based on the dimeric structure (green and cyan) of the A. vinelandii NifL LOV domain (Protein Data Bank code 2GJ3). FAD binding to each NifL PAS domain is shown. The N-terminal α-helix (residues Ser⁶¹¹–Ser⁶²²) of PAS2 predicted to be involved in dimerization is indicated. C, PAS2 residues Glu⁶⁶⁶ and Leu⁶⁷⁴, corresponding to the NifL FAD binding residues Thr⁷⁸ and Leu⁸⁶. D, potential PAS2 residues involved in ligand binding were assessed using the β-galactosidase assay with different ligands. The N-terminal PAS2 α-helix (residues Ser⁶¹¹–Ser⁶²²) predicted to be involved in dimerization, as seen in Fig. 4B, was also evaluated with the same assay. E, the modeled PAS2 was superimposed onto toluene-bound PAS1. The PAS2 residues Tyr⁶⁹¹ and Ala⁷⁰³ were analyzed with the corresponding PAS1 residues Ile¹¹⁴ and Val¹²⁶, which are responsible for toluene binding.